Lipoproteins in vivo are mainly classified according to its specific gravity into four groups, i.e. high-density lipoproteins (hereinafter abbreviated as HDL), low-density lipoproteins (hereinafter abbreviated as LDL), very-low-density lipoproteins (hereinafter abbreviated as VLDL), and chylomicrons (hereinafter abbreviated as CM).
Lipid composition and types of apoprotein differ for each lipoprotein, and thus their function in vivo differs significantly with one another. Further, there is also an intermediate-density lipoprotein (hereinafter abbreviated as IDL) being intermediate between VLDL and LDL, as a lipoprotein which is generated during metabolism of VLDL to LDL, while it is classified as LDL in a broad sense.
At present, when screening for diagnosing arterial sclerosis in the field of clinical laboratory test, total cholesterol, total triglyceride, HDL cholesterol, apolipoprotein AI, apolipoprotein B, etc. are generally measured. Recently, LDL cholesterol which is said to be highly associated with arteriosclerosis formation is frequently measured instead of total cholesterol. On the other hand, there are many patients showing coronary atherosclerosis lesion while their LDL cholesterol in blood is not high, and it is said that a condition showing adipositas, hypertriglyceridemia, hypertension, and hyperglycemia in combination is a very dangerous clinical condition that might develop coronary artery diseases. Further, it is also reported that the triglycerides level in LDL in blood is associated with arterial diseases more than LDL cholesterol.
General methods for measuring triglycerides in LDL include a method of fractionating and collecting lipoproteins by ultracentrifugation, precipitation method or using immunoreactions, and measuring triglycerides contained therein. However, all the fractionating methods require time and cost, and are very complicated.
As a method measurable with an autoanalyzer without carrying out fractionation, a method comprising removing triglycerides in all lipoproteins except LDL in the first step, and measuring triglycerides in the remaining LDL in the second step is known (patent references 1 and 2). However, there is a problem that in a sample with high triglycerides in VLDL or CM, triglycerides cannot be completely removed in the first step, and the reaction is carried over to the second step, causing a positive impact.
Further, as a method measurable with an autoanalyzer without carrying out operations for fractionation, a method for measuring triglycerides contained in IDL, which is classified as LDL in a broad sense, is known (patent reference 3). However, the method does not measure triglycerides contained in normal LDL, and therefore the method is not appropriate for measuring triglycerides in LDL.
Further, as a method measurable with an autoanalyzer without carrying out fractionation, a method is known for measuring triglycerides in LDL characterized in that an enzyme reaction is carried out in the presence of a block copolymer of propylene oxide and ethylene oxide, particularly a triblock copolymer of polyoxypropylene and polyoxyethylene (patent reference 4).
Patent Reference 1: WO 00/43537
Patent Reference 2: WO 2004/087945
Patent Reference 3: WO 00/60112
Patent Reference 4: Published Japanese Translation of PCT International Publication No. 2002-508519